プローブDNA結合粒子によるHIV-DNAの特異的な濃縮及びそのPCR検出  [in Japanese] Prior Enrichment of HIV-DNA with Probe-DNA Particles for an Efficient PCR Diagnosis  [in Japanese]

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Author(s)

Abstract

probe-DNAを結合したラテックス粒子を用いて, HIV-DNAを特異的に濃縮してから, PCR法で増幅することにより, 極めて高感度なHIV-DNAの検出法を確立した. 数十m1中に数コピーないし数十コピーの標的DNAが存在する系においては, 通常のPCRではその検出が難しい. 本方法によれば, 粒子表面のprobe-DNAが極めて高い効率で標的DNAとハイブリダイゼーションすることができ, また, 捕捉されたHIV-DNAを粒子から解離せず, 粒子ごとにPCR反応で増幅することができた.<BR>モデル検体系に於いて本方法の検出感度は18コピー/30m1であった. また, HIVキャリアーの血液1部を正常血液29部, 49部, 99部と混合したプール血液系について, 通常のPCRではいずれも検出できなかったが, 本方法の濃縮後PCRでは, 全て陽性の結果が得られた. 従って, 本方法は, PCR法による輸血用血液のHIVスクリーニングへの道を拓くものと期待できる.

PCR mediated detection of HIV-DNA has been widely used. However, compared with traditional immunological diagnoses, the extraction of DNA is a laborious and time consuming step. We have developed a procedure for the efficient isolation and concentration of HIV-DNA from cell lysates. We report here a novel method by which one can recover HIV-DNA in a small volume (-50μl) of solution from a large volume of crude cell lysate which contains as few as several copies. The method uses the specific hybridization of HIV-DNA to HIV probe-DNA particles. This prior enrichment augmented the sensitivity in the detection of HIV-DNA by PCR, and allows us to make a diagnosis even if the specimen contained an extremely low copy number of HIV-DNA molecules in a large volume, which would have otherwise resulted in false-negative data with the conventional extraction method. The method also enables the examination of 100 individual blood specimens in a combined form. Thus, the application of the present enrichment procedure with HIV probe-DNA particles should reduce the labor and cost of HIV diagnosis, since the HIV positive samples represent a very minor group of people among specimens subjected to clinical laboratory tests and, particularly, among blood samples voluntarily donated to be used for transfusions.

Journal

  • Kansenshogaku Zasshi

    Kansenshogaku Zasshi 69(9), 957-962, 1995-09-20

    The Japanese Association for Infectious Diseases

References:  8

Codes

  • NII Article ID (NAID)
    10008727072
  • NII NACSIS-CAT ID (NCID)
    AN00047715
  • Text Lang
    JPN
  • Article Type
    ART
  • ISSN
    03875911
  • NDL Article ID
    3644806
  • NDL Source Classification
    SC191(感染症)
  • NDL Source Classification
    ZS9(科学技術--医学--病理学・微生物学・寄生虫学・感染・免疫学・血清学・アレルギー)
  • NDL Call No.
    Z19-193
  • Data Source
    CJP  NDL  J-STAGE 
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